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Oncology - Basic: Basic ScienceAntibodies and Larger Molecules |
1 Stanford University Medical Center, Molecular Imaging Program at Stanford (MIPS), Palo Alto, CA
386
Objectives: Site-specifically label a 3-helix scaffold based protein (ZEGFR1907) binding to epidermal growth factor receptor (EGFR) with 64Cu, investigate its in vitro cell uptake and binding specificity, study the 64Cu labeled ZEGFR1907 (64Cu-DOTA-ZEGFR1907) in vivo tumor imaging ability.
Methods: DOTA-ZEGFR1907 was made through solid phase synthesis of the protein Cys-ZEGFR1907 followed by the site specific conjugation with a DOTA derivative. The bioconjugate was then labeled with 64Cu in sodium acetate buffer (pH=5.0) at room temperature. Cell uptake test was performed with A431 cell line and pre-incubation with/without unlabeled DOTA-ZEGFR1907. 64Cu-DOTA-ZEGFR1907 (30-50 µCi) was injected into A431 tumor bearing nude mice through tail-vein for microPET imaging and bio-distribution studies. In vitro analysis of EGFR levels in tumor, liver and blood was studied by western blot analysis using anti-human EGFR antibody.
Results: 64Cu-DOTA- ZEGFR1907 displayed a moderate specific activity (30 µCi/µg). In vitro cell uptake of 64Cu-DOTA-ZEGFR1907 was high (~25%) and specific. In vivo microPET imaging showed fast tumor targeting, high tumor accumulation (>10 %ID/g at 4 h p.i.) and good contrast of 64Cu-DOTA-ZEGFR1907. Bio-distribution studies demonstrated that 64Cu-DOTA-ZEGFR1907 had high tumor, blood, liver and kidney uptakes, while blood level was dropped significantly 24 h p.i.. It was also observed high EGFR expression levels in blood, liver and tumor as tested by western blot analysis.
Conclusions: 64Cu-DOTA-ZEGFR1907 can provide high contrast, receptor specific PET imaging of EGFR positive tumors. High expression of EGFR in blood doesn’t interfere with 64Cu-DOTA-ZEGFR1907 accumulation in tumors.
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